This invention relates to the identification of new genes coding for cellular constituents involved in the regulation of angiogenesis. The invention relates more specifically to a process for the identification of these genes. The invention also relates to the use of the factors coded by the identified genes for the clinical study of the angiogenesis process, for the diagnosis and treatment of pathologies linked to this process as well as for pharmacological, pharmacogenomic or drug identification trials.
Angiogenesis is the fundamental process by which new blood vessels are formed. This process is essential in numerous normal physiological phenomena such as reproduction development and cicatrization. In these normal biological phenomena, angiogenesis is under strict control, i.e., it is triggered during a short period of time, several days, then completely inhibited. However, numerous pathologies are linked to invasive, uncontrolled angiogenesis. Arthritis, for example, is a pathology caused by damage to cartilage by invasive neovessels. In diabetic retinopathy, the invasion of the retina by neovessels leads to blindness in the patients. Neovascularization of the ocular apparatus is a major cause of blindness and such neovascularization is responsible for around twenty different diseases of the eye. Finally, the growth and metastasis of tumors is directly linked to neovascularization and is dependent on angiogenesis. The tumor stimulates the growth of the neovessels for its own growth. Moreover, these neovessels provide escape routes allowing the tumors to reach the blood circulation and cause metastases at remote sites such as the liver, lungs and bones.
In other pathologies such as the cardiovascular diseases, the diseases of the peripheral arteries as well as the vascular and cerebral lesions, angiogenesis can present an important therapeutic base. In fact, the promotion of angiogenesis in the damaged locations can lead to formation of sanguineous neovessels that are lateral and alternative to the damaged vessels, thereby providing blood and, thus, oxygen and other nutritive and biological factors necessary for the survival of the tissues involved.
Formation of neovessels by endothelial cells involves migration, growth and differentiation of endothelial cells. Regulation of these biological phenomena is directly linked to the genetic expression. With regard to angiogenesis, a constantly growing number of studies show that the regulation of angiogenesis is implemented via an equilibrium among the factors acting directly on the endothelial cell. These factors can be stimulating factors, on the one hand, such as VEGF, FGFs, IL-8, HGF/SF, PDGF, etc. They can also be inhibitory factors such as IL-10, IL-12, gro-xcex1 and xcex2, platelet factor 4, angiostatin, human chondrocyte-derived inhibitor, thrombospondin, leukemia inhibitory factor, etc. (Jensen, Surg. Neural., 1998, 49, 189-195; Tamatani et al., Carcinogenesis, 1999, 20, 957-962; Tanaka et al., Cancer Res., 1998, 58, 3362-3369; Ghe et al., Cancer Res., 1997, 57, 3733-3740; Kawahara et al., Hepatology, 1998, 28, 1512-1517; Chandhuni et al., Cancer Res., 1997, 57, 1814-1819; Jendraschak and Sage, Semin. Cancer Biol., 1996, 7, 139-146; Majewski et al., J. Invest. Dermatol., 1996, 106, 1114-1119).
The control of angiogenesis, thus, represents a strategic axis both for fundamental research to improve comprehension of the numerous pathological phenomena linked to angiogenesis, as well as a foundation for the development of new therapies for treating pathologies linked to angiogenesis.
This invention relates to a process for identifying genes coding for cellular constituents involved in regulating angiogenesis including culturing endothelial cells on an extracellular matrix protein according to at least four different types of conditions: a reference condition, an angiogenesis promoting condition, an angiogenesis inhibiting condition, and a control condition; isolating messenger RNAs stemming from cells cultured according to the different conditions; and comparing at the qualitative and/or quantitative level, different messenger RNA populations to identify messenger RNAs stemming exclusively or in a particularly elevated quantity from cell cultures under conditions stimulating and/or inhibiting angiogenesis, the messenger RNAs corresponding to the genes coding for the cellular constituents involved in regulating angiogenesis.
This invention also relates to a pharmaceutical composition for diagnosing and/or treating pathologies linked to angiogenesis including a gene involved in regulating angiogenesis identified by the process for identifying genes coding for cellular constituents involved in regulating angiogenesis including culturing endothelial cells on an extracellular matrix protein according to at least four different types of conditions: a reference condition, an angiogenesis promoting condition, an angiogenesis inhibiting condition, and a control condition; isolating messenger RNAs stemming from cells cultured according to the different conditions; and comparing at the qualitative and/or quantitative level, different messenger RNA populations to identify messenger RNAs stemming exclusively or in a particularly elevated quantity from cell cultures under conditions stimulating and/or inhibiting angiogenesis, the messenger RNAs corresponding to the genes coding for the cellular constituents involved in regulating angiogenesis.
This invention further relates to a laboratory kit for diagnosing and/or treating pathologies linked to angiogenesis, including products and reagents for culturing endothelial cells on an extracellular matrix protein in combination with one or more angiogenic and anti-angiogenic factors.